Histone modifications have been implicated in stem cell maintenance and differentiation. We have analyzed genome-wide changes in gene expression and histone modifications during differentiation of multipotent human primary hematopoietic stem/progenitor cells (HSCs/HPCs) into erythrocyte precursors. Our data indicate that H3K4me1, H3K9me1 and H3K27me1 associate with enhancers of differentiation genes prior to their activation and correlate with basal expression, suggesting that these mono-methylations are involved in the maintenance of activation potential required for differentiation. In addition, although the majority of genes associated with both H3K4me3 and H3K27me3 in HSCs/HPCs become silent and lose H3K4me3 after differentiation, those that lose H3K27me3 and become activated after differentiation are associated with increased levels of H2A.Z, H3K4me1, H3K9me1, H4K20me1 and RNA polymerase II in HSCs/HPCs. Thus, our results suggest that gene expression changes during differentiation are programmed by chromatin modifications present at the HSC/HPC stage and we provide a resource for enhancer and promoter identification.